LOCUS       Exported                7979 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pPtPBR1, complete sequence.
ACCESSION   KX523201
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7979)
  AUTHORS   Diner RE, Bielinski VA, Dupont CL, Allen AE, Weyman PD.
  TITLE     Refinement of the Diatom Episome Maintenance Sequence and 
            Improvement of Conjugation-Based DNA Delivery Methods
  JOURNAL   Front Bioeng Biotechnol 4, 65 (2016)
  PUBMED    27551676
REFERENCE   2  (bases 1 to 7979)
  AUTHORS   Diner RE, Weyman PD.
  TITLE     Direct Submission
  JOURNAL   Submitted (07-JUL-2016) Scripps Institution of Oceanography, 
            University of California San Diego, 9500 Gilman Drive, La Jolla, CA 
            92093, USA
REFERENCE   3  (bases 1 to 7979)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..7979
                     /organism="Cloning vector pPtPBR1"
                     /mol_type="other DNA"
                     /db_xref="taxon:1897460"
     promoter        10..38
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     regulatory      10..15
                     /regulatory_class="minus_35_signal"
                     /note="tet (P2) promoter"
     regulatory      33..38
                     /regulatory_class="minus_10_signal"
                     /note="tet (P2) promoter"
     regulatory      complement(44..49)
                     /regulatory_class="minus_10_signal"
                     /note="P1 promoter"
     regulatory      complement(64..75)
                     /regulatory_class="minus_35_signal"
                     /note="P1 promoter -35 sequence region"
     CDS             86..1276
                     /codon_start=1
                     /product="tetracycline efflux protein class C"
                     /function="confers resistance to tetracyclin"
                     /label=tetracycline efflux protein class C
                     /protein_id="AOI27793.1"
                     /translation="MKSNNALIVILGTVTLDAVGIGLVMPVLPGLLRDIVHSDSIASHY
                     GVLLALYALMQFLCAPVLGALSDRFGRRPVLLASLLGATIDYAIMATTPVLWILYAGRI
                     VAGITGATGAVAGAYIADITDGEDRARHFGLMSACFGVGMVAGPVAGGLLGAISLHAPF
                     LAAAVLNGLNLLLGCFLMQESHKGERRPMPLRAFNPVSSFRWARGMTIVAALMTVFFIM
                     QLVGQVPAALWVIFGEDRFRWSATMIGLSLAVFGILHALAQAFVTGPATKRFGEKQAII
                     AGMAADALGYVLLAFATRGWMAFPIMILLASGGIGMPALQAMLSRQVDDDHQGQLQGSL
                     AALTSLTSIIGPLIVTAIYAASASTWNGLAWIVGAALYLVCLPALRRGAWSRATST"
     regulatory      1905..1909
                     /regulatory_class="ribosome_binding_site"
                     /note="rop RBS"
     CDS             1915..2106
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(2337..2708)
                     /codon_start=1
                     /gene="traJ"
                     /product="oriT-recognizing protein"
                     /label=traJ
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     oriT            2741..2850
                     /note="incP origin of transfer"
     primer_bind     3127..3143
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        3167..3185
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     CDS             3889..4263
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     misc_feature    4569..5072
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     misc_feature    4569..4685
                     /label=yeast centromere
                     /note="yeast centromere"
     rep_origin      4686..5073
                     /label=yeast origin of replication
                     /note="yeast origin of replication"
     promoter        5073..5259
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     CDS             5260..5922
                     /codon_start=1
                     /gene="S. cerevisiae HIS3"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=HIS3
                     /note="yeast auxotrophic marker"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALGAVRGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHF
                     LESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     rep_origin      complement(6152..6740)
                     /direction=LEFT
                     /note="/note=pMB1 origin of replication (RNAII -35 to 
                     RNA/DNA switch point)"
     misc_RNA        complement(6152..6704)
                     /label=RNAII transcript
                     /note="RNAII transcript"
     regulatory      6558..6563
                     /regulatory_class="minus_35_signal"
                     /note="RNAI promoter; TTGAAG"
     regulatory      6580..6585
                     /regulatory_class="minus_10_signal"
                     /note="RNAI promoter; GCTACA"
     misc_RNA        6594..6701
                     /label=RNAI transcript
                     /note="RNAI transcript"
     regulatory      complement(6714..6719)
                     /regulatory_class="minus_10_signal"
                     /note="RNAII promoter; CGTAAT"
     regulatory      complement(6735..6740)
                     /regulatory_class="minus_35_signal"
                     /note="RNAII promoter; TTGAGA"
     CDS             complement(6911..7771)
                     /codon_start=1
                     /gene="Bla"
                     /product="beta-lactamase"
                     /function="confers resistance to ampicillin"
                     /label=Bla
                     /protein_id="AOI27794.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            complement(6911..7771)
                     /gene="Bla"
                     /label=Bla
     sig_peptide     complement(7703..7771)
                     /gene="Bla"
                     /note="required for secretion to the periplasm; cleaved off
                     to form the mature beta-lactamase protein"
     promoter        complement(7772..7876)
                     /gene="bla"
                     /label=AmpR promoter
     regulatory      complement(7779..7783)
                     /regulatory_class="ribosome_binding_site"
                     /note="bla RBS"
     regulatory      complement(7815..7820)
                     /regulatory_class="minus_10_signal"
                     /note="bla (P3) promoter; GAGACA"
     regulatory      complement(7836..7841)
                     /regulatory_class="minus_35_signal"
                     /note="bla (P3) promoter; TTCAAA"